Recombinant anti-mouse TIGIT antibody

Recombinant murinized TIGIT antibody for in vivo use

Anti-mTIGIT-mIgG2a InvivoFit™ is a recombinant murinized anti-mouse (m)TIGIT monoclonal antibody (mAb) featuring the variable region of the previously described 10A7 clone [1] and a mouse (m)IgG2a constant region.

InvivoGen’s engineered Anti-mTIGIT-mIgG2a InvivoFit™ antibody

TIGIT (T cell immunoglobulin and ITIM domain) is a cell surface inhibitory immune checkpoint [1]. It is specifically expressed on immune cells including natural killer (NK) cells, activated and memory T cells, as well as regulatory T cells (Tregs) [2].

The 10A7 mAb has been used to block TIGIT signaling in Tregs and chronically stimulated CD8+ T cells [3, 4]. Using recombinant technology, the original 10A7 hamster IgG2a constant region has been replaced with a murine IgG2a format which mediates potent cytotoxic functions [5]. This murinization also allows for reduced immunogenicity of the administered mAb and risks of fatal hypersensitivity reactions [6-8].
Anti-mTIGIT-mIgG2a is provided in an InvivoFit™ grade, a high-quality standard specifically adapted to in vivo studies.

Key features of Anti-mTIGIT-mIgG2a InvivoFit™:

• Derives from the 10A7 clone, Hamster IgG2a
• Features the mIgG2a isotype (constant region)
• Filter-sterilized (0.2 µm), endotoxin level < 1 EU/mg
• Suitable for parental delivery in mice (azide-free)
• Low aggregation < 5%
• Produced in animal-free facilities and defined media

Anti-mTIGIT-mIgG2a InvivoFit™ is produced in Chinese hamster ovary (CHO) cells, purified by affinity chromatography with protein A, and its binding to mTIGIT is validated using flow cytometry.

References:

1. Yu X. et al, 2009. The surface protein TIGIT suppresses T cell activation by promoting the generation of mature immunoregulatory dendritic cells. Nat Immunol. 10(1):48-57.
2. Chauvin J.M. & Zarour H.M., 2020. TIGIT in cancer immunotherapy. J. Immunother. Cancer 8:e000957.
3. Jonhston R.J., et al,  2014. The immunoreceptor TIGIT regulates antitumor and antiviral CD8(+) T cell effector function. Cancer Cell. 26(6):923-937.
4. Joller, N. et al,  2014. Treg Cells Expressing the Coinhibitory Molecule TIGIT Selectively Inhibit Proinflammatory Th1 and Th17 Cell Responses. Immunity. 40(4):569-581.
5. Nimmerjahn F. & Ravetch J.V., 2005. Divergent immunoglobulin g subclass activity through selective Fc receptor binding. Science. 310:1510-2.
6. Mall C. et al., 2016. Repeated PD-1/PD-L1 monoclonal antibody administration induces fatal xenogenic hypersensitivity reactions in a murine model of breast cancer. Onco Immunol. 5(2):e1075114.
7. Murphy, J.T.et al,  2014. Anaphylaxis caused by repetitive doses of a GITR agonist monoclonal antibody in mice. Blood. 123:2172-80.
8. Belmar N.A. et al,  2017. Murinization and H chain isotype matching of Anti-GITR antibody DTA-1 reduces immunogenicity-mediated anaphylaxis in C57BL/6 mice. J Immunol. 198:4502-4512.

Figures

Cell surface staining of murine TIGIT using Anti-mTIGIT-mIgG2a InvivoFit™ mAb.
~5x10⁵ EL4 (parental) orEL4-mTIGIT cells were incubated with 2 µg of Anti‑mTIGIT‑mIgG2a InvivoFit™ mAb or an isotype control for 1h at 4°C. Cells were then washed and incubated with 0.25 µg of goat anti-mIgG2a coupled to PE for 1h at 4°C. Cell surface staining was analyzed by flow cytometry.

Specifications

Specificity: Targets cells expressing murine TIGIT

Formulation: Lyophilized from 0.2 μm filtered solution in 150 mM sodium chloride, 20 mM sodium phosphate buffer with 5% saccharose

Clonality: Monoclonal antibody

Isotype: Murine IgG2a, Kappa

Control: mIgG2a  InvivoFit™ isotype control

Source: CHO cells

Purity: Purified by affinity chromatography with protein A

Applications: Flow cytometry; in vivo depletion; ELISA

Quality control:

• Binding of Anti-mTIGIT-mIgG2a InvivoFit™ to mouse TIGIT has been confirmed using Flow cytometry
• The complete sequence of the antibody construct has been verified
• < 5% aggregates (confirmed by size exclusion chromatography)
• Endotoxin level <1 EU/mg (determined by the LAL assay)

Contents

Anti-mTIGIT-mIgG2a InvivoFit™ is filter-sterilized (0.2 µm), endotoxin-free, azide-free, and lyophilized.  

This product is available in two pack sizes:

• mtigit-mab10-1​: 1 mg
• mtigit-mab10-10​: 10 mg

 The product is shipped at room temperature.

 Store lyophilized antibody at -20 °C.

 Lyophilized product is stable for at least 1 year

 Avoid repeated freeze-thaw cycles.

InvivoFit™

InvivoFit™ is a high-quality standard specifically adapted for in vivo studies. InvivoFit™ products are filter-sterilized (0.2 µm) and filled under strict aseptic conditions in a clean room. The level of bacterial contaminants (endotoxins and lipoproteins) in each lot is verified using a LAL assay and a TLR2 and TLR4 reporter assay.

Details

TIGIT Background:

TIGIT (T cell immunoglobulin and ITIM domain) is a 27 KDa cell surface immunoreceptor described as an inhibitory immune checkpoint [1]. TIGIT is specifically expressed on immune cells including, Natural Killer (NK) cells, activated and memory T cells, as well as regulatory T cells (Tregs) [2]. TIGIT binds to CD155 (PVR) and CD112 (PVRL2, nectin-2) expressed on antigen-presenting cells (APCs), T cells, and a variety of non-hematopoietic cells, including tumor cells [1, 2]. Of note, TIGIT competes with CD226 (also known as DNAM-1) and CD96 (also known as Tactile) for the same ligands [1, 2]. TIGIT interaction with its ligand triggers the phosphorylation of its cytoplasmic ITIM motif, and subsequent inhibition of the NF-κB, P13K, and MAPK pathways. As a result, TIGIT inhibits NK and CD8+ T cell effector functions [1-3]. In Tregs, TIGIT ligation triggers the production of immunosuppressive molecules such as IL-10 [4]. TIGIT is a target for monoclonal antibodies in cancer treatments [2].

1. Yu X. et al, 2009. The surface protein TIGIT suppresses T cell activation by promoting the generation of mature immunoregulatory dendritic cells. Nat Immunol. 10(1):48-57.
2. Chauvin J.M. & Zarour H.M., 2020. TIGIT in cancer immunotherapy. J. Immunother. Cancer 8:e000957.
3. Jonhston R.J., et al,  2014. The immunoreceptor TIGIT regulates antitumor and antiviral CD8(+) T cell effector function. Cancer Cell. 26(6):923-937.
4. Joller, N. et al,  2014. Treg Cells Expressing the Coinhibitory Molecule TIGIT Selectively Inhibit Proinflammatory Th1 and Th17 Cell Responses. Immunity. 40(4):569-581.